Bacterial populations associated with the human body play important roles in both health and disease. The objective of this research is to understand how altered patterns of bacterial colonization impact outcomes after major traumatic injury. This proposal is designed as an ancillary study to the ADAPT trial, a NIH-funded observational study of best practices in the care of pediatric traumatic brain injury (TBI). The work is an extension of a pilot study of critically ill children that documented widespread derangements of the microbiome, which in some cases were linked to nosocomial infections. We will use next-generation DNA sequencing to resolve population level bacterial colonization patterns in 50 patients with severe TBI. We will profile colonization patterns using high-throughput 16S rRNA sequencing of stool samples, skin swabs, tongue swabs, tracheal aspirates, and samples of cerebrospinal fluid collected twice weekly after admission to the PICU. A panel of 4 microbiome analyses will be performed to profile the microbiota for each individual patient, and for the entire study population. We will use group-based trajectory analysis to track changes in diversity, presence of dominant nosocomial pathogens, and loss of microbial site-specificity across body sites. Alterations in the microbiota of TBI patients will be compared to colonization patterns in control populations of adults and children (Aim 1). We will use multiple statistical approaches including a hierarchical mixed effect linear model to characterize how the ICU microbiome is impacted by clinical variables, including antibiotic exposure, enteral nutrition, and contamination from the ICU room environment (Aim 2). Finally, we will utilize regression models to test the hypothesis that specific derangements in the microbiome increase the odds of adverse clinical outcomes including nosocomial infection, prolonged hospital stay, death, and poor neurologic function at 6 months (Aim 3). Improved understanding of post-injury colonization patterns could translate to improved outcomes for trauma victims by suggesting more effective patient care strategies, e.g. modification of the microbiome to prevent infection. More broadly, this research will uncover aspects of colonization dynamics with implications for a diverse range of critically ill patients, including both children nd adults.